Flow control system for diagnostic assay system

ABSTRACT

A disposable cartridge for mitigating cross-contamination of fluid sample reagents. The disposable cartridge includes a cartridge body defining a syringe barrel having an barrel port operative to inject and withdraw assay fluids in response to the displacement of a syringe plunger. Furthermore, the disposable cartridge includes a rotor defining a plurality of assay chambers in fluid communication with the barrel port through one of a plurality of rotor ports disposed about the periphery of the rotor. Finally, the disposable cartridge includes a flow control system between the barrel and rotor ports which prevents cross-contamination of fluid sample reagents from one assay chamber to another assay chamber.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuing application of U.S. National patentapplication Ser. No. 16/302,303, entitled “FLOW CONTROL SYSTEM FORDIAGNOSTIC ASSAY SYSTEM” and filed Nov. 16, 2018, which is a 371National Application of PCT International Application No.:PCT/US2017/032904, filed May 16, 2017, which further claims priority toa U.S. Provisional Patent Application Ser. No. 62/337,423 filed May 17,2016 entitled “Modified Desalting Column” and a second U.S. ProvisionalPatent Application Ser. No. 62/337,446 filed May 17, 2016 entitled“Multi-Chamber Rotating Valve and Cartridge” The contents of theaforementioned applications are hereby incorporated by reference intheir entirety.

This application also relates to U.S. patent application Ser. No.15/157,584 filed May 18, 2016 entitled “Method and System for SamplePreparation” which is a continuation of U.S. Non-Provisional patentapplication Ser. No. 14/056,603, filed Oct. 17, 2013, now U.S. Pat. No.9,347,086, which claims priority to U.S. Provisional Patent ApplicationSer. No. 61/715,003, filed Oct. 17, 2012, which is acontinuation-in-part of U.S. patent application Ser. No. 12/785,864,filed May 24, 2010, now U.S. Pat. No. 8,663,918, which claims priorityto U.S. Provisional Patent Application Ser. No. 61/180,494, filed May22, 2009, and which is also a continuation-in-part of U.S. patentapplication Ser. No. 12/754,205, filed Apr. 5, 2010, now U.S. Pat. No.8,716,006, which claims priority to U.S. Provisional Patent ApplicationSer. No. 61/166,519, filed Apr. 3, 2009. The contents of theaforementioned applications are hereby incorporated by reference intheir entirety.

TECHNICAL FIELD

This invention relates to a disposable cartridge for use in combinationwith a diagnostic assay system which performs RNA polymerase-DNAanalysis of a biological sample. The assay system drives a rotor about arotational axis as a syringe plunger injects and withdraws sample fluidsinto and out of the disposable cartridge. Embodiments of a disposablecartridge are disclosed including variations which facilitate flow,repeatability, reliability, admixture, and preparation of the assayfluids.

BACKGROUND

There is continuing interest to improve testing methodologies,facilitate collection and decrease the time associated with clinicallaboratories. Particular testing requires that a sample be disrupted toextract nucleic acid molecules such as DNA or RNA.

The number of diagnostic tests performed annually has increasedexponentially in the past decade. The use of molecular diagnostics andgene sequencing in research and medical diagnostics is also rapidlygrowing. For example, DNA testing has also exploded in view of thegrowing interest in establishing and tracking the medical history and/orancestry of a family. Many, if not all of these assays, could benefitfrom a rapid sample preparation process that is easy to use, requires nooperator intervention, is cost effective and is sensitive to a smallsample size.

Sample collection and preparation is a major cost component ofconducting real-time Polymerase Chain Reaction (PCR), gene sequencingand hybridization testing. In addition to cost, delays can lead to thespread of infectious diseases, where time is a critical component to itscontainment/abatement. In addition to delaying the test results, suchactivities divert much-needed skilled resources from the laboratory tothe lower-skilled activities associated with proper collection, storageand delivery.

For example, a portable molecular diagnostic system could be operated byminimally trained personnel (such as described in US 2014/0099646 A1)and have value with regard to disease surveillance. However, theadoption of such portable systems can be limited/constrained by currentmethods of sample collection, which require trained personnel to permitsafe and effective handling of blood/food/biological samples foranalysis. Other limitations relate to: (i) the ability ofinjected/withdrawn fluids to properly flow, (ii) manufacturability,(iii) cross-contamination of assay fluids which may influence theveracity of test results, (iv) proper admixture of assay fluids toproduce reliable test results, and (v) the ability or inability tointroduce catalysts to speed the time of reaction,

A need, therefore, exists for an improved disposable cartridge for usein combination with a portable molecular diagnostic/assay system whichfacilitates/enables the use of minimally-trained personnel, hands-offoperation (once initiated), repeatable/reliable test results acrossmultiple assay samples (e.g., blood, food, other biological samples) andan ability to cost effectively manufacture the disposable cartridge forthe diagnostic assay system.

SUMMARY

The present disclosure relates to a variety of disposable cartridgeconfigurations for a portable molecular diagnostic/assay system.

In one embodiment, a filtration column assembly is provided for use incombination with a disposable cartridge of a diagnostic assay system.The filtration column assembly includes a column matrix materialconfigured to filter a fluid sample, a tubular column configured tosealably engage a filtration chamber of the disposable cartridge and acap configured to be inserted into an end of the tubular column anddefine a passageway to direct the sample fluid from the second end ofthe tubular column into a collection cavity disposed adjacent thefiltration chamber. The tubular column defines: (i) a column cavity forreceiving the column matrix material, (ii) a first end having an openingfor receiving the fluid sample and configured to retain the columnmatrix material, and (iii) a second end, receiving the fluid directingcap, and having an opening to dispense a filtered fluid sample from thecolumn cavity.

In another embodiment, a disposable cartridge is provided for mitigatingcross-contamination of fluid sample reagents. This embodiment includes acartridge body defining a syringe barrel having an barrel port operativeto inject and withdraw assay fluids in response to the displacement of asyringe plunger. Furthermore, the disposable cartridge includes a rotordefining a plurality of assay chambers disposed in fluid communicationwith the barrel port through one of a plurality of rotor ports disposedabout the periphery of the rotor. Finally, the disposable cartridgeincludes a flow control system between the barrel and rotor ports whichprevents cross-contamination of fluid sample reagents from one assaychamber to another assay chamber.

BRIEF DESCRIPTION OF THE DRAWINGS

The present invention is disclosed with reference to the accompanyingdrawings, wherein:

FIG. 1 is a perspective view of a portable diagnostic assay systemoperative to accept one of a plurality of disposable cartridgesconfigured to test fluid samples of collected blood/food/biologicalsamples.

FIG. 2 is an exploded perspective view of one of the disposablecartridges configured to test a blood/food/biological sample.

FIG. 3 is a top view of the one of the disposable cartridgesillustrating a variety of assay chambers including a central assaychamber, one of which contains an assay chemical suitable to breakdownthe fluid sample to detect a particular attribute of the tested fluidsample.

FIG. 4 is a bottom view of the disposable cartridge shown in FIG. 3illustrating a variety of channels operative to move at least a portionof the fluid sample from one chamber to another the purpose ofperforming multiple operations on the fluid sample.

FIG. 5 is an exploded perspective view of the disposable cartridgeincluding a filtration column assembly comprising a column matrixmaterial, a tubular column receiving and retaining the column matrixmaterial, and a fluid directing cap configured to restrict the volume ofa filtered fluid sample while maintaining a pressurized flow path.

FIG. 6 is a cross-sectional view taken substantially along line 6-6 ofFIG. 5 wherein a plurality of ports are depicted in a common plane ofthe cartridge rotor.

FIG. 7 is a cross-sectional view taken substantially along line 7-7 ofFIG. 6 depicting the filtration column disposed in a filtration chamberof the disposable assay cartridge.

FIG. 8 is an isolated perspective view of the filtration columndepicting a fluid guide operative to direct the filtered fluid sampleinto a collection chamber.

FIG. 9 is an exploded perspective view of the filtration columnincluding the column matrix material, the tubular column for receivingthe column matrix material, the fluid directing cap and a retentionfilter melted, welded, or otherwise attached to the lower end of thetubular column.

FIG. 10 is an exploded, isolated, perspective view of the fluiddirecting cap depicting the underside surface of the cap, including aretention filter disposed in combination with an annular rim of thefluid directing cap.

FIG. 11 is a schematic side sectional view of the filtration columnincluding the column matrix material for trapping/filtering smallmolecule materials while allowing the passage of large molecules.

FIG. 12 is a cross-sectional view taken substantially along line 12-12of FIG. 5 wherein a plurality of rotor ports are depicted in a commonplane along with the syringe barrel and a barrel port of the cartridgebody.

FIG. 13 depicts an exploded perspective view of the disposable cartridgeassembly including a compliant over-mold disposed between the rotor andthe cartridge body.

FIG. 14 is a cross-sectional view of the disposable cartridge takensubstantially along line 14-14 of FIG. 12 depicting an internal view ofthe compliant over-mold to show the location of several compliantopenings in the over-mold.

FIG. 15 is a cross-sectional view of the disposable cartridge takensubstantially along line 15-15 of FIG. 14 depicting an X-shaped valvedisposed in a compliant over-mold for preventing backflow contaminationfrom one assay chamber to another.

FIG. 16 is an enlarged plan view of the X-shaped valve depicting thesize differential between the opening of the X-shaped valve and thediameter of the barrel port of the cartridge body.

FIG. 17 is a cross-sectional view of the disposable cartridge takensubstantially along line 17-17 of FIG. 12 depicting an internal view ofthe compliant over-mold to show the location of several compliantopenings in the over-mold.

FIG. 18 is a cross-sectional view of the disposable cartridge takensubstantially along line 18-18 of FIG. 17 depicting a flap valvedisposed in the compliant over-mold for preventing backflowcontamination from one assay chamber to another assay chamber.

FIG. 19 is an enlarged plan view of the flap valve depicting the sizedifferential between the opening of the flap valve and the diameter ofthe barrel port of the cartridge body.

FIG. 20 is a sectional view through the disposable cartridge depicting avortex generator disposed downstream of a rotor port to facilitatemixture of the assay fluids within the respective chamber of the rotor.

FIG. 21 is an enlarged, broken-away side view of the vortex generatorshown in FIG. 20 .

FIG. 22 is an bottom view of the rotor depicting an enlarged channelwhich may be heated to accelerate reagent reactions within the channel.

FIG. 23 depicts a bottom view of the rotor depicting segmented channelsfor performing various PCR reactions therein.

FIGS. 24A-E depict a series of primer interactions wherein a well isempty in FIG. 24A, a buffer suspends or re-suspends the primer in FIG.24B, the buffer is purged in FIG. 24C, rehydrated primers are sealed inthe respective suspension wells in FIG. 24D, and the PCR product isdiffused in FIG. 24E.

FIGS. 25 a-25 d are sectional views of the disposable cartridgeillustrating various alternative embodiments for preventingcross-contamination from one disposable cartridge to another when usinga common diagnostic assay test device wherein FIG. 25 a depicts adisposable shaft, FIG. 25 b depicts a series of compliant washersdisposed within the syringe barrel to limit exposure of a permanentshaft of the diagnostic assay device to the assay fluids, FIG. 25 cshows a bellows diaphragm to fully contain the fluids within the syringebarrel, and FIG. 25 d illustrates a primary working plunger disposed incombination with a secondary plunger for preventing the permanent shaftfrom exposure to the contaminating assay fluids.

Corresponding reference characters indicate corresponding partsthroughout the several views. The examples set out herein illustrateseveral embodiments of the invention but should not be construed aslimiting the scope of the invention in any manner.

DETAILED DESCRIPTION

A disposable cartridge is described for use in a portable/automatedassay system such as that described in commonly-owned, co-pending U.S.patent application Ser. No. 15/157,584 filed May 18, 2016 entitled“Method and System for Sample Preparation” which is hereby included byreference in its entirety. While the principal utility for thedisposable cartridge includes DNA testing, the disposable cartridge maybe used in be used to detect any of a variety of diseases which may befound in either a blood, food or biological specimen. For example, blooddiagnostic cartridges may be dedicated cartridges useful for detectinghepatitis, autoimmune deficiency syndrome (AIDS/HIV), diabetes,leukemia, graves, lupus, multiple myeloma, etc., just naming a smallfraction of the various blood borne diseases that the portable/automatedassay system may be configured to detect. Food diagnostic cartridges maybe used to detect salmonella, e-coli, Staphylococcus aureus ordysentery. Diagnostic cartridges may also be used to test samples frominsects and animals to detect diseases such as malaria, encephalitis andthe west nile virus, to name but a few.

More specifically, and referring to FIGS. 1 and 2 , a portable assaysystem 10 receives any one of a variety of disposable assay cartridges20, each selectively configured for detecting a particular attribute ofa fluid sample, each attribute potentially providing a marker for ablood, food or biological (animal borne) disease. The portable assaysystem 10 includes one or more linear and rotary actuators operative tomove fluids into, and out of, various compartments or chambers of thedisposable assay cartridge 20 for the purpose of identifying ordetecting a fluid attribute. More specifically, a signal processor 12,i.e., a PC board, controls a rotary actuator (not shown) of the portableassay system 10 so as to align one of a variety of ports 18P, disposedabout a cylindrical rotor 18, with a syringe barrel 22B of a stationarycartridge body 22. The processor 14 controls a linear actuator 24, todisplace a plunger shaft 26 so as to develop pressure i.e., positive ornegative (vacuum) in the syringe barrel 22. That is, the plunger shaft26 displaces an elastomer plunger 28 within the syringe 22 to move andor admix fluids contained in one or more of the chambers 30, 32.

The disposable cartridge 20 provides an automated process for preparingthe fluid sample for analysis and/or performing the fluid sampleanalysis. The sample preparation process allows for disruption of cells,sizing of DNA and RNA, and concentration/clean-up of the material foranalysis. More specifically, the sample preparation process of theinstant disclosure prepares fragments of DNA and RNA in a size range ofbetween about 100 and 10,000 base pairs. The chambers can be used todeliver the reagents necessary for end-repair and kinase treatment.Enzymes may be stored dry and rehydrated in the disposable cartridge 20,or added to the disposable cartridge 20, just prior to use. Theimplementation of a rotary actuator allows for a single plunger 26, 28to draw and dispense fluid samples without the need for a complex systemof valves to open and close at various times. This greatly reducespotential for leaks and failure of the device compared to conventionalsystems. Finally, it will also be appreciated that the system greatlydiminishes the potential for human error.

In FIGS. 3 and 4 , the cylindrical rotor 18 includes a central chamber30 and a plurality of assay chambers 32, 34 surrounded, and separatedby, one or more radial or circumferential walls. In the describedembodiment, the central chamber 30 receives the fluid sample while thesurrounding chambers 32, 34 contain a premeasured assay chemical orreagent for the purpose of detecting an attribute of the fluid sample.The chemical or reagents may be initially dry and rehydrated immediatelyprior to conducting a test. Some of the chambers 32, 34 may be open toallow the introduction of an assay chemical while an assay procedure isunderway or in-process. The chambers 30, 32, 34 are disposed in fluidcommunication, i.e., from one of the ports 18P to one of the chambers30, 32, 34, by channels 40, 42 molded along a bottom panel 44, i.e.,along underside surface of the rotor 18. For example, a first port 18P,corresponding to aperture 42, may be in fluid communication with thecentral chamber 30, via aperture 50.

Filtration Cartridge

During development of the disposable cartridge, and as the inventorsacquired an appreciation for, and understanding of, the fluid dynamicsinvolved with respect to injecting, dispensing and withdrawing the assayfluids, they discovered that surface tension between components cansignificantly impact fluid flow from one chamber 32 to another chamber34. As a consequence, they learned that the properties of surfacetension can detrimental or advantageous to fluid flow. For example,surface tension between a film cover 60 (see FIG. 5 ), which encloses orencapsulates the assay fluids of the various chambers 30, 32, 34, 36,and an upper end of a filtration column assembly 100, can prevent theflow of assay fluids from the filtration chamber 34 to an adjacentcollection chamber 36. The following discloses embodiments of afiltration column assembly 100 which facilitates fluid flow from thefiltration chamber 34 to the collection chamber 36.

In FIG. 5 , a novel filtration column assembly 100 comprises: (i) acolumn matrix material 112 configured to filter a fluid sample; (ii) atubular column 114 having a column cavity for receiving/retaining thecolumn matrix material and configured to sealably engage a filtrationchamber 34 of the disposable cartridge 20, and (iii) a fluid directingcap 116, configured to be detachably mounted to one end of the columnand defining a passageway (not viewable in FIG. 5 ) configured to directa filtered fluid sample from the tubular column 114 into the collectionchamber 36. In another embodiment of the novel filtration columnassembly 100, the cap 116 is configured to restrict the volume of thefiltered fluid sample collected above the column matrix material 112while maintaining a pressurized flow path.

In FIG. 6 , a cross-sectional view through the key-hole shapedfiltration and collection chambers 34, 36 reveals a wall 38 separatingthe chambers 34, 38. FIG. 3 also depicts a separating wall 38 which isabout one-half (½) the wall height of the filtration chamber 34. In theview shown, the filtration chamber 34 includes a plurality of raisedsurfaces or ridges 74 to facilitate flow of the fluid sample from a port74 through the cylindrical wall 76 of the disposable cartridge 20, i.e.,of the rotor 18. That is, assay fluid will be injected into the port 74which will, in turn flow between the ridges 74, and into a bottom end ofthe tubular column 114,

Inasmuch as the intervening wall 38, i.e., the wall separating thefiltration and collection chambers 34, 36, is less than the full wallheight of the filtration chamber 34, it will be appreciated that, whenfluid in the tubular column 114 reaches this level or height, the samplefluid will accumulate in the filtration chamber 34 and flow over thewall 38 between the chambers 34, 36. To prevent flow from taking thispath or direction, i.e., over the wall 38, without travelling throughthe length of the tubular column 114, the bottom portion 124 of thetubular column 114 is configured to sealably engage the surroundingfiltration and collection chamber walls 68. This forces pressurizedassay fluid to flow upward through the length of the tubular column 114.

The column matrix material 112 is a filter material which is operativeto secure, trap or chemically bond, a select material suspended orflowing with, a carrier fluid, i.e., water, lysis fluid, etc. In theillustrated embodiments, the column matrix material 112 has transitionedfrom a dehydrated condition, i.e., shown in phantom in FIG. 9 , to ahydrated condition, i.e., shown in solid lines in FIGS. 5, 7, 9 and 11 .Initially, the column matrix material 112 is significantly smaller thanthe column cavity 122 and is retained therein by a screening materialdisposed at each end of the tubular column 112. More specifically, afirst filter material 132 is melted, welded or otherwise affixed to thefirst end of the tubular column 114 while a second filter material 134is melted, welded or otherwise affixed to the removable cap 116 of thefiltration column 100, i.e., when the cap 116 is inserted in the secondend of the tubular column 114. As such, the first filter material 132retains the column matrix material by closing-off the first opening 124in the bottom end of the column 114 while the second filter material 134retains the column matrix material 112 by closing-off the second end ofopening 126 in the tubular column 114.

Upon contact with the fluid sample, the column matrix material 112 ishydrated to fill the width of the tubular column 114. Furthermore, thecolumn matrix material 112 grows to a prescribed length necessary toremove the target molecular material from the fluid sample. As fluidpasses through the column matrix material 112, it traps small moleculematerials in the matrix while allowing large molecule materials to pass.In the described embodiment, the small molecule material is sodiumchloride (i.e., salt), however, the fluid dynamics described herein areapplicable to any filtration material requiring a particular length ofmatrix material to remove a select molecule. For example, the columnmatrix material 112 may remove materials from a group comprising, butnot limited to: phosphates, sodium and polysaccharides. In the describedembodiment, the large molecule material may be a deoxyribonucleic acid(DNA) molecule. It is this large molecule material which will ultimatelybe deposited in the collection chamber 36 and screened for testing.

It should be appreciated that the filter material 132, 134 may be anyconventional screening material which allows molecules of a particularsize to pass. In the described embodiment, the filter materials 132, 134allow both large and small molecules to pass. Accordingly, the filtermaterials 132, 134 do not remove molecules from the fluid assay sample,but merely function as a convenient solution to retain the both thedehydrated and hydrated column matrix material in the tubular column114. Furthermore, the filter materials 132, 134 allow the passage ofmolecules larger than those trapped by the column matrix material 114.

The cap 116 is configured to be inserted into the second or upper end126 of the tubular column 112, functionally retains the other end of thecolumn matrix material 114, and defines a passageway configured todirect a filtered fluid sample from the tubular column 114 into thecollection chamber 36. Alternatively, or additionally, the cap 116 maybe configured to restrict the volume of the filtered fluid samplecollected above the column matrix material 112 and maintain apressurized flow path.

The cap 116 includes a substantially planar cover 136, an annular rim138 projecting orthogonally from the plane of the cover 136, and a fluidguide 142 also projecting from the cover 136 and defining a fluid pathor passageway 150 from the upper portion of the column matrix material112 to the collection chamber 36. The annular rim 138 of the cap 116 isconfigured to be inserted into the upper end, i.e., into the upperopening 126, of the tubular column 114 and includes an effluent opening146 (best seen in FIG. 10 ) aligned with a transverse opening 148 in thewall of the tubular column 114. More specifically, the fluid guide 142of the cap 116 is configured to receive a filtered fluid sample from thealigned openings 144, 146 to direct the filtered fluid sample into thecollection chamber 36 of the disposable cartridge 20.

In FIG. 11 , a schematic of the filtration column 100 is depicted withthe cap 116 inserted in the upper end 126 of the tubular column 114. Aseluded to supra, the cap 116 reduces the volume of filtered assay fluidwhich may collect on the top of the column matrix material 112. As such,the cap 116 retains sufficient head pressure, and creates a pressurizedpassageway 150 which avoids the difficulties associated with the surfacetension between the upper end of the filtration column 100 and a filmcover which encapsulates the various cavities and chambers 30, 32, 34,36 of the disposable cartridge 20.

Flow Control System

While the previous section disclosed an embodiment relating to onechamber for filtration and another for collection of a filtered assayfluid, the following section relates to improvements pertaining to flowcontrol between the syringe barrel 22B and the plurality of ports 16disposed about the periphery of the rotor 18. During a period ofdevelopment, understanding and discovery, the inventors learned that themanufacture of the disposable cartridge 20, and in particular, themanufacture of the syringe barrel 22B and the rotor 18, presentedcertain challenges that could only be addressed by a novel compliantover-mold 200 disposed between the rotor 18 and the stationary cartridgebody 22. Before discussing the compliant over-mold 200, it will helpfulto understand some other considerations pertaining to the need for thecompliant over-mold 200.

Various methods for manufacturing the rotor and cartridge body 22 wereconsidered in the early stages of the cartridge development. It will beappreciated that for the cartridge to be disposable, (i.e., used onlyonce), it must be extremely inexpensive to fabricate, i.e., to satisfythe margins necessary for profitability. On the other hand, both therotor and cartridge body are intricate, i.e., requiring a multiplicityof very narrow apertures 18P and channels 40, 42 (shown in FIGS. 3 and 4). One manufacturing approach considered by the inventors involved acombination of molding and machining steps. For example, the rotor 18and cartridge body 22 may be independently molded, and subsequentlymachined to produce the many intricate rotor and syringes ports 18P,22P. Furthermore, this approach facilitates the fabrication of therequisite narrow ports 18P, 22P, which enables surface tension developedin the ports 18P, 22P to prevent backflow contamination of the assayfluids. While this approach provides the desired port dimensions, thesubsequent machining operation is far too costly for a diagnostic systemwhich employs disposable cartridges 20.

Another approach involved injection molding which significantly reducesmanufacturing costs, however, this method also has certain limitationsrelating to the dimension/diameter of the rotor and syringe ports 18P,22P. More specifically, the molding pins used to fabricate the ports18P, 22P must maintain a certain threshold dimension to prevent themolding pins from failing or fracturing during the injection moldingprocess. As such, the requisite pin size for fabricating the ports 18P,22P is significantly larger than the optimum port dimension forpreventing backflow contamination. As such, the port size which must bemaintained cannot take advantage of the properties of surface tension toprevent backflow contamination. Consequently, a need arose for afabrication method which employs injection molding as the principlefabrication technique (to keep costs to a minimum) while producing therequisite port size without resorting to more expensive manufacturingmethods.

In FIGS. 12 and 13 , disposable cartridge 20 comprises a rotor 18,cartridge body 22, and a flow control system operative to preventcross-contamination of fluid sample reagents from one assay chamber toanother assay chamber. As described supra, the rotor 18 comprises aplurality of assay chambers 30, 34 rotatable about an axis 18A androtationally mounted to the cartridge body 22. Furthermore, the rotor 18defines a peripheral surface, i.e., a cylindrical surface 18S, having aplurality of ports 18P disposed about the surface 18S and extendingthrough the wall 18W which defines the peripheral surface 18S. Thecartridge body 22, on the other hand, comprises a syringe barrel 22Boperative to inject and withdraw assay fluids in response to axialdisplacement of a syringe plunger 28 disposed within the syringe barrel22B. Both the rotor 18 and cartridge body 22 includes ports 18P and 22P,respectively, which are fabricated using a conventional injectionmolding process. Accordingly, the port size is limited by the pinrestraints of an injection molding process.

To combat the difficulties associated with cross-contamination of fluidsample reagents, a moldable, compliant valve, or elastomer over-mold200, was interposed between the rotor 18 and cartridge body 22. In FIGS.12-16 , the elastomer or compliant over-mold 200 defines at least onecompliant opening 204 having a maximum opening dimension which issmaller than the dimension of the rotor ports 18P. Furthermore, thecompliant openings 204 has a maximum opening dimension which is smallerthan the dimension of the barrel port 22P. In the described embodimentthe average opening dimension of the barrel port 22P is about 1.3 mmwhereas the opening dimension of the compliant opening is less thanabout 0.6 mm and preferably less than about 0.3 mm. Operationally, thecompliant opening 204 is configured to: (i) enlarge when fluid pressureis applied in response to axial movement of the plunger 28 duringinjection and (ii) diminish in size when fluid pressure is reduced.

In one embodiment, depicted in FIGS. 14-16 , the compliant opening 204includes intersecting cuts 212 configured to cross the opening of thebarrel port 22P. In this embodiment, the X-shaped opening has flexiblecorner segments 204A, 204B which may flap or bend so as to produce alarger opening, i.e., to permit a larger flow of assay fluid through thecompliant opening 204. Consequently, the corner segments 204A, 204B ofthe compliant opening 204 function as a valve, i.e., opening to permit agreater flow rate in response to positive pressurization and closing inthe absence of the positive pressurization. It will also be appreciatedthat the segments 204A, 204B may flex in the opposite direction to allowfluid to be withdrawn from an assay chamber 30, 32, 34. Accordingly, thecompliant X-shaped opening can also function as a two-way valve, i.e.,opening in one direction to allow flow in that direction and in anotherdirection to allow flow in the opposite direction. To further increasethe flow rate across the intersecting cuts 204, a portion of thecross-over region, i.e., the portion closest or proximal to thecross-over cuts can be removed or eliminated to facilitate flow. In thedescribed embodiment, the portion of the cross-over region 212 which isremoved is less than about 0.5 mm and in another embodiment the region212 is less than about 0.3 mm.

In another embodiment, depicted in FIGS. 17-19 , the compliant opening208 may comprise a flap 210 mounted to an edge of the opening 208 by aflexible elastomer hinge 214. In this embodiment, the compliant opening208 includes a frustum-shaped opening 218 while the flap 210 includes anedge which is complimentary-shaped to seat in the frustum-shaped opening218. Similar to the previous embodiment, the compliant opening 208 has amaximum opening dimension which is smaller than the dimension of therotor ports 18P. Furthermore, the compliant opening 208 has a maximumopening dimension which is smaller than the dimension of the barrel port22P. Both the rotor and barrel ports 18P, 22P are shown in dashed orphantom lines in FIGS. 17-19 .

Operationally, the flap 210 of the compliant opening 208 is unseated asthe pressure within the syringe barrel increases to inject assay fluidinto the rotor 16 through one of the ports 18P. The increased pressurecauses the flap 210 to pivot about the hinge axis to dispense the assayfluid into an assay chamber 32, 34, 36. Once this step is completed, thepressure is withdrawn such that the flap 210 closes and is reseated intothe frustum-shaped opening. Next, the signal processor provides a signalto rotate the rotor to a new rotational position. The flap 210 of thecompliant opening functions to prevent backflow of the recentlydeposited assay fluid into the syringe barrel 22B. As such, the X-shapedcompliant opening 204, and the hinged flap 208 contained in theelastomer over-mold 200, prevent assay fluids from being wicked or drawn(should the syringe barrel 22 retain a small negative pressure or pocketof positive pressure) into the barrel port 22P. Accordingly, theover-mold valves 204, 208 ensure that the test results will not betainted and will be accurate.

In yet another embodiment shown in FIGS. 12 and 13 , the ports of theflow control system may be disposed on different geometric planes.Alternatively or additionally, ports having compatible reagents may bein fluid communication with one syringe barrel/plunger while portshaving incompatible reagents may be in fluid communication with adifferent syringe barrel/plunder so as to separate/isolate incompatiblereagents from each other. For example, reagents of one type may beinjected by one of the syringe barrels/plungers 22B-1, 28-1 whilereagents of another type may be injected by another one of the syringebarrels/3081061 plungers 22B, 28-2. In FIG. 13 , a backside surface ofthe elastomer over-mold 200 is shown depicting two compliant openings224, 228. One of the X-shaped compliant openings 224 is disposedproximal the bottom surface or plane of the rotor 18 and is fed by afirst syringe barrel 22B-1 deployed on one plane of the disposablecartridge 20. A second X-shaped compliant opening 228 is fed by a secondsyringe barrel 22B-1 deployed on a second plane of the disposablecartridge 20. In the described embodiment, the port 224 disposed in oneplane is separated or spaced apart from the port 228 disposed in anotherplane by a threshold or prescribed vertical distance.

In another embodiment of the disclosure, at least one of the rotor ports18P includes a high viscosity gel disposed in the bore of the respectiveport 18P. The high viscosity is injected into at least one of the rotorports 18P such that the gel extends the full length of the port, i.e.,on average about 1.5 mm.

Operationally the gel is displaced under pressure to facilitate thetransfer of fluid sample reagents from one assay chamber to another. Inthe described embodiment, at least one of the rotor ports 18P define afluid volume which is less than about 15 microliters to mitigateback-flow of a fluid sample reagent.

Co-Molded or Dual Material Rotor for Enhanced Thermal and ConformalProperties

In another embodiment of the disclosure, the rotor 18 comprisesdifferent materials to enhance the thermal and conformal properties ofthe disposable cartridge 20 Depending upon material compatibility, therotor 18 may be molded in segments and subsequently joined/welded toform a complete rotor 18. By fabricating the rotor 18 employing at leasttwo different materials, e.g., one segment having conductive propertiesand another segment fabricated from a high modulus material (having highstrain properties), the rotor 18 can provide enhanced performance. Forexample, a lower portion of the rotor 18 can be fabricated usingconductive materials to function as a heat sink. As such, a heatingelement (not shown in the drawing) can deliver heat to various chambers32, 34, 36 and channels 40, 42 to accelerate reagent reactions andimprove the performance of the disposable cartridge 20.

The rotor segments can be fabricated using a thermally conductiveplastic or a thermally conductive elastomer. Both materials havesuperior thermal properties to standard polypropylene while the additionof elastomer has added conformal properties. Lastly, inasmuch as theupper segment of the rotor may comprise a material having low thermalconductivity, this segment will have insulating properties to retainheat in regions where it provides the most benefit.

Enhanced Mixing

One of the requirements of the disposable cartridge 20 is the admixtureof reagent fluids in the various chambers 30, 32, 34, 36 to ensure acomplete, thorough and reliable result. While the diagnostic assaysystem 10 may include shakers, mixers and vibration inducing actuators,one of the easiest structures to accomplish mixing in a chamber includesa spinner, vortex generator or flow disruptor. FIGS. 20 and 21 depictanother embodiment of the disposable cartridge wherein a vortexgenerator 300 is disposed above a port 18P extending through the bottompanel of the rotor 18. As assay fluid is injected into the port 18P, thefluid immediately encounters a disruptor in the fluid flow. The vortexgenerator 300 generates mixing vortices causing the injected fluid tomix with a fluid, e.g., a lysis buffer, in the chamber 32 of the rotor18.

Sumps, Tapered Floors and Rounded Corners

In another embodiment and referring to FIG. 22 , the configuration ofthe rotor 18 and its chambers 30, 32, 34 and 36 may be configured tobetter extract all the contents from a reservoir. As such, sump areasmay be created, corners of the walls may be rounded and the bottom panel44 may be pitched or inclined in a direction toward a port 54 projectingupwardly through the bottom panel 44 of the rotor 18.

Heated Channels

In another embodiment and referring to FIG. 22 , a large channel region52 may be located on the bottom 44 of the rotor 18 and configured to besufficiently large to retain and entire sample and lysing buffermixture. As such, the high volume channel 52 may be positioned over aheating element (not shown) and integrated with a film disposed over thebottom channels 40, 42, and 52, to accelerate the reagent reactionsoccurring in the mixture.

Isolated Multi-zoned Multiplexed PCR using a Single Buffer

In yet another embodiment and referring to FIG. 23 , one or morechannels 40, 42 will be constructed in a segmented design in which PCRprimers may be dispensed, dried and stored in individual segments 400. Asubsequent coating may be applied to further encapsulate the driedprimers 410. The coating serves two functions: (1) to preserve andprotect the primers and (2) to prevent premature rehydration of theprimers when the channel 420 is filled with a buffer material. Concernsrelating to lateral diffusion may be minimized by the low primerdiffusion rate, spot to spot distance (e.g., 3 to 5 mm.) and the use ofnarrow channels separating the spotted regions (choke point).

In another embodiment depicted in FIGS. 24A-E, isolation of the PCRregions can be achieved by utilizing small wells 500 created in a bottomfilm using a laminated film processing technique. The micro wells 500would contain the dried primers 510 and other desired components. InFIG. 24A, upon filling the micro wells 500 with a common buffer, themicro wells 500 may be filled in a step (ii) to re-suspend the primers510 in FIG. 24B. A secondary, non-miscible fluid such as mineral oil maybe added in a step (iii) to cap the micro wells 500 in FIG. 24C. Withthe micro wells sealed, in step (iv), with the mineral oil, the PCRprocess can be performed in in FIG. 24D. Extraction of the fluid in step(v) would first involve replacing the mineral oil with a suitableaqueous buffer. Due the large concentration gradient, a large quantityof the PCR product would then diffuse into the buffer in FIG. 24E.

To prevent the primer 510 from spreading during the loading phase, anencapsulant may be used. The encapsulant may be water soluble,semi-water soluble or temperature sensitive in order to preventimmediate rehydration of the primers. Upon filling, the encapsulant willslowly dissolve and eventually allow for the primers to be re-suspendedinto the buffer. A temperature sensitive encapsulant would maintain itsintegrity until a critical temperature is reached, wherein it is brokendown allowing the primers to re-suspend.

Syringe Isolation and Containment

While previous embodiments involved the prevention ofcross-contamination from chamber to chamber, the possibility forcross-contamination can occur from one disposable cartridge to anotherdisposable cartridge. For example, the possibility exists that thesyringe shaft 26, which is part of the portable diagnostic assay system10, may be contaminated by a previously used disposable cartridge 20.That is, the shaft 26 which actuates the plunger 28 may be contaminatedby assay materials in the syringe barrel 22, i.e., as the shaft wipesagainst the barrel opening for receiving the shaft 26.

FIGS. 26 a-26 e depict various configurations of the syringe barrel 22for preventing cross-contamination between cartridges 20. During anormal syringe actuation, the shaft which drives the plunger 28 isexposed to the outside environment (see FIG. 26 a ). During this state,it is in theory, possible for trace reagent residue and particulates tobe left exposed due to insufficient plunger sealing/scraping and thusrisking contamination on the syringe shaft or other areas. In oneembodiment, shown in FIG. 26 a , a disposable shaft 600 detachably mateswith the plunger 28 at one end and removeably mounts to a permanentshaft (not shown) within the portable diagnostic assay system 10. Thisminimizes the risk of syringe shaft contamination as it is discardedalong with the cartridge 20.

In another embodiment, illustrated in FIG. 26 b , a disposable shaft 600passed through a series of elastomer flaps/baffles 610 which function asmultiple gaskets. The flexible nature of the elastomer allows the shaft600 to operate while maintaining intimate contact with the shaftreducing the possibility of exposure.

In another embodiment, depicted in FIG. 26 c , a disposable shaft 600 isconnected to one end of a flexible bellows 620 which, in turn, mounts atits other end of the syringe barrel 28. As the shaft 600 extends andretract, the bellows 620 expands and collapses This configurationcompletely isolates the syringe shaft 600 from the internal environmentof the syringe barrel 28.

In yet another embodiment, shown in FIG. 26 d , the disposable shaft 600connects to a second plunger 630 disposed a threshold distance X fromthe primary or working plunger 28. The disposable shaft 600, workingplunger 28 and secondary plunger 630 is inserted into a syringe barrel22 which is elongated by the same threshold distance between theplungers 28, 630. The primary plunger 600 is used as a traditionalplunging mechanism and is responsible for moving the fluid into thecartridge. The secondary plunger 630 functions as a containment deviceand is spaced from the primary plunger 28 such that secondary orcontainment plunger 630 never passes into the working area (or stroke)of the primary plunger 28. This prevents contaminants from beingconveyed to the secondary or containment plunger 630. A permanent shaft650 extends beyond the elongated syringe barrel 22 where it is mated tothe linear actuator or control motor a syringe control motor.

While the invention has been described with reference to particularembodiments, it will be understood by those skilled in the art thatvarious changes may be made and equivalents may be substituted forelements thereof without departing from the scope of the invention. Inaddition, many modifications may be made to adapt a particular situationor material to the teachings of the invention without departing from thescope of the invention.

Therefore, it is intended that the invention not be limited to theparticular embodiments disclosed as the best mode contemplated forcarrying out this invention, but that the invention will include allembodiments falling within the scope and spirit of the appended claims.

What is claimed is:
 1. A disposable cartridge for use in a diagnosticassay system, comprising: a cartridge body defining a syringe operativeto inject and withdraw assay fluids through a barrel port; a rotorcomprising a plurality of assay chambers and mounted for rotation to thecartridge body about an axis, the rotor defining a plurality of rotorports disposed about a peripheral surface, each rotor port disposed influid communication with at least one assay chamber of the plurality ofassay chambers; and a compliant over-mold interposing the cartridge bodyand the rotor and defining at least one compliant opening interposingthe barrel port and one of the plurality of rotor ports.
 2. Thedisposable cartridge of claim 1 wherein the compliant opening includesan aperture having a dimension smaller than a dimension of an aperturedefined by the barrel port.
 3. The disposable cartridge of claim 1wherein the compliant opening includes an aperture having a dimensionsmaller than a dimension of an aperture defined by the barrel port and adimension of each of the rotor ports and wherein the compliant openingincreases when fluid pressure is applied and decreases when fluidpressure is reduced.
 4. The disposable cartridge of claim 2 wherein thecompliant opening includes a flap covering the opening.
 5. Thedisposable cartridge of claim 2 wherein the compliant opening includesintersecting cuts disposed through the over-mold.
 6. The disposablecartridge of claim 2 wherein the compliant opening includes intersectingcuts disposed through the over-mold, wherein the intersecting cutsdefine a cross-over region and wherein a portion of the cross-overregion is removed to facilitate fluid flow through the cross-overregion.
 7. The disposable cartridge of claim 4 wherein the flap includesan elastomer hinge configured to open and close the flap.
 8. Thedisposable cartridge of claim 6 wherein the portion of the cross-overregion is less than about 0.5 mm.
 9. The disposable cartridge of claim 6wherein the portion of the cross-over region is less than about 0.3 mm.10. A disposable cartridge for use with a diagnostic assay system,comprising: a cartridge body defining a syringe operative to inject andwithdraw assay through a barrel port; a rotor comprising a plurality ofassay chambers and mounted for rotation to the cartridge body about anaxis, the rotor defining a plurality of rotor ports disposed about aperipheral surface, each rotor port disposed in fluid communication withat least one of the plurality of assay chambers; a flow control systembetween the barrel port and the rotor ports configured to preventcross-contamination of fluid sample reagents from one assay chamber toanother assay chamber.
 11. The disposable cartridge of claim 10 whereinthe flow control system comprises at least two flow control portswherein the at least two flow control ports of the flow control systemare disposed in a common plane normal to the axis.
 12. The disposablecartridge of claim 10 wherein the flow control system comprises at leasttwo flow control wherein the at least two flow control ports of the flowcontrol system are disposed in different planes normal to the axis. 13.The disposable cartridge of claim 12 wherein the at least two flowcontrol ports of the flow control system associated with one plane arespaced-apart from the at least two flow control ports associated withanother plane by a prescribed vertical distance.
 14. The disposablecartridge of claim 10 wherein at least one of the rotor ports include ahigh viscosity gel disposed in a bore of the port, the high viscositygel containing the fluid sample reagents their respective chambers priorto use.
 15. The disposable cartridge of claim 10 wherein at least one ofthe rotor ports include a high viscosity gel disposed in a bore of therotor port, the high viscosity gel being displaced under pressure toenable a transfer of fluid sample reagents from one assay chamber toanother assay chamber.
 16. The disposable cartridge of claim 10 whereinat least one of the rotor ports define a fluid volume which is less thanabout 15 microliters to prevent back-flow of a fluid sample reagent. 17.The disposable cartridge of claim 10 wherein the flow control systemincludes an elastomer over-mold interposing the cartridge body and atleast a portion of the peripheral surface of the rotor, the elastomerover-mold having at least one compliant opening corresponding to thebarrel port.
 18. A method for preventing cross-contamination of fluidsample reagents in a diagnostic assay system, comprising the steps of:configuring a cartridge body with a syringe operative to withdraw andinject assay fluids through a barrel port of the syringe and into aplurality of assay chambers of a cartridge rotor, the cartridge rotordisposed within and rotatable about an axis of the cartridge body, thecartridge rotor having a peripheral surface defining a plurality ofrotor ports, and each of the plurality of rotor ports disposed in fluidcommunication with at least one of the assay chambers; interposing acompliant over-mold between the cartridge body and the rotor, thecompliant over-mold defining a first compliant opening aligned with afirst rotor port of the plurality of rotor ports and defining a secondcompliant opening aligned with a second rotor port of the plurality ofrotor ports, selectively aligning the barrel port with the firstcompliant opening of the compliant over-mold to withdraw an assay fluidfrom a first assay chamber of the plurality assay chambers into thesyringe of the cartridge body; and selectively aligning the barrel portwith the second compliant opening of the compliant over-mold to injectan assay fluid into a second assay chamber of the plurality of assaychambers from the syringe of the cartridge body wherein the compliantopening prevents cross-contamination of fluids drawn from the firstassay chamber of the plurality of assay chambers of the cartridge bodyand into the second assay chamber of the plurality of assay chambers ofthe cartridge body.
 19. The method of claim 18 wherein each of the firstand second compliant openings have a dimension smaller than a dimensionof an aperture defined by the barrel port.
 20. The method of claim 18wherein each of the first and second compliant openings have a dimensionsmaller than a dimension of an aperture defined by the barrel port and adimension of each of the plurality of rotor ports and wherein each ofthe first and second compliant openings increase when a fluid pressureis applied and decrease when a fluid pressure is reduced.
 21. The methodof claim 18 wherein each of the first and second compliant openingsinclude a flap covering the opening.
 22. The method of claim 18 whereineach of the first and second compliant openings include intersectingcuts disposed through the over-mold.
 23. The method of claim 18 whereineach of the first and second compliant openings include intersectingcuts disposed through the over-mold, wherein the intersecting cutsdefine a cross-over region and wherein a portion of the cross-overregion is removed to facilitate fluid flow through the cross-overregion.
 24. The method of claim 21 wherein the flap includes anelastomer hinge configured to open and close the flap.